Induction of breast cancers depends on ovarian steroids. However, at diagnosis, 2/3 of tumors have switched, from growth control by steroid agonists, to control by peptide growth factors. Hypothesis: progesterone primes breast cancers for the proliferative effects of growth factors, which then down-regulate progesterone receptors (PR) creating a negative feedback loop. The remaining 1/3 of steroid hormone-dependent tumors respond well to antagonists but acquire resistance as they progress. Hypothesis: acquired resistance to antagonists is associated with inappropriate expression of their agonist effects. Aim 1. Resistance to Agonists. Is steroid resistance biochemically linked to upregulation of growth factor signaling? Using stable cell lines expressing A- or B-isoforms of PR, or mutant PR, we will study priming effects of progesterone and the antiprogestin RU486: on acquired sensitivity to proliferative effects of EGF; on upregulation of MAPK and STAT signaling; on MAPK phosphorylation of PR which targets them for ubiquitylation and downregulation; on transcriptional synergism at p21 and c-fos promoters. These studies will define mechanisms for cross-talk between PR and EGF in the cytoplasm and nucleus. Aim 2. Acquired Resistance to Anatagonists. Are the agonist effects of mixed antagonists upregulated by recruitment of regulatory proteins to antagonist-occupied PR? Three novel proteins have been isolated in an antagonist-biased yeast two hybrid screen. ORF number 93 has multiple NR boxes and at least 3 TPR domains. We will clone it, and ask whether it acts as a scaffold to assemble PR in a multi-protein complex with EGF signaling molecules; is a coregulator; or assembles PR into a repressive transcription complex. The mechanisms of two other novel proteins will be assessed. These studies will describe the function of antagonist-occupied receptors as governed by three new receptor- interacting proteins. Aim 3. Acquired Resistance to Antagonists: Breast Cancers. Does the transcriptional coactivator to corepressor ratio determine outcome to tamoxifen? Tumors from tamoxifen responsive or resistant patients will be measured for transcript expression of the coactivators L7/SPA and SRC-1, and the corepressors N-CoR and SMRT, in relation to treatment response. We will show that mechanisms of SMRT repression are novel. Other studies address EGF signaling molecules and ORF number 93. These studies will document the relationships between coregulators and tamoxifen resistance, and demonstrate that antagonist-occupied steroid receptors recruit factors inappropriately. Overall, these studies will explain hormone-resistance mechanisms in breast cancers.